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Publisher Summary The chapter focuses on the application of liquid chromatography (LC) tandem mass spectrometry (MS) (LC-MS-MS)-based approaches to identifying protein targets of modifications and mapping the modifications at the level of amino acid sequence. This chapter describes (1) the tandem MS (MS-MS) analysis of modified peptides and the impact of different adduct chemistries on peptide ion fragmentation, (2) a new MS-MS data analysis tool called scoring algorithm for spectral analysis (SALSA), (3) the application of SALSA to sequence-specific mapping of modifications to proteins, and (4) the complementary relationship of SALSA to other data analysis algorithms for MS-MS data. The integration of sequest and SALSA provides a powerful approach to proteome characterization by LC-MS-MS. The different peptide modifications confer three types of characteristics on MS-MS spectra: (1) product ions derived from adduct moieties, (2) neutral or charged losses arising from adduct-specific fragmentations, and (3) shifts in b- or y-ion signals along the m/z axis. SALSA evaluates MS-MS spectra for specific user-defined features, including product ions at specific m/z values, neutral or charged losses from singly or doubly charged precursors, and ion pairs or series. The ability of SALSA to identify unanticipated variants is a distinct advantage over other data analysis tools. Sequest and related programs can correlate the MS-MS data of modified peptides with database sequences if the user specifies the nature of the modification and the amino acid modified. |
